The critically ill patient frequently develops a complex disease spectrum that may include acute respiratory distress syndrome, systemic inflammatory response syndrome, septic shock and multiple organ dysfunction syndrome (MODS). Attempts at developing effective therapies for sepsis/septic shock and MODS have proven to be exceedingly difficult. We hypothesize that the systemic inflammatory response to sepsis can be attenuated or aborted by using an alpha7 nicotinic receptor agonist to activate the cholinergic anti- inflammatory pathway. This pathway is part of a reflex arc that modulates immune responses by acting at the spleen to suppress production of pro-inflammatory cytokines by macrophages. The neural components of this pathway comprise preganglionic vagal efferent nerves (cholinergic) and their target postganglionic sympathetic neurons (noradrenergic) that innervate the spleen. Recent evidence suggests that local release of norepinephrine activates a cholinergic phenotype in splenic T cells, but in situ expression of cholinergic markers in the spleen has not been explored. Suppression of cytokine production by macrophages is mediated by alpha7 nicotinic receptors. Activated T cells may be the source of acetylcholine (ACh) that stimulates these receptors. However, macrophages can also acquire a cholinergic phenotype and might be an additional source for ACh. Furthermore, splenic monocytes also produce inflammatory cytokines and could be additional targets for ACh. Thus, there are many unknowns regarding mechanisms involved in converting the neural signal into an anti-inflammatory/anti-sepsis response. The clinical significance of the cholinergic anti-inflammatory pathway is amplified by evidence that its benefits appear to reach beyond the spleen, but the full extent of the systemic response is yet to be established. This project will address these gaps in knowledge by using a clinically relevant murine model of polymicrobial sepsis (CLP) and evaluating histochemical, biochemical, and therapeutic responses evoked by treatment with a drug (GTS-21) that activates the cholinergic anti-inflammatory pathway by stimulation of alpha7 nicotinic receptors. Aim 1 will employ biochemical and immunohistochemical methods to: 1) identify and characterize splenocytes that make and/or respond to ACh and 2) evaluate effects of GTS-21 to increase ACh concentration in the spleen and suppress production of inflammatory cytokines by splenocytes. Aim 2 will evaluate the therapeutic response of septic mice to treatment with GTS-21. Different dosing regimens will be used to evaluate the efficacy of GTS-21 at preventing, attenuating, or reversing sepsis-induced cardiac dysfunction, pulmonary pathology, and immunosuppression. Results from this study will expand our understanding of the mechanisms and therapeutic potential of the cholinergic anti-inflammatory pathway.